• Evolution of cytokine production capacity in ancient and modern European populations.

      Domínguez-Andrés, Jorge; Kuijpers, Yunus; Bakker, Olivier B; Jaeger, Martin; Xu, Cheng-Jian; Van der Meer, Jos Wm; Jakobsson, Mattias; Bertranpetit, Jaume; Joosten, Leo Ab; Li, Yang; et al. (eLife Sciences Publications, 2021-09-07)
      As our ancestors migrated throughout different continents, natural selection increased the presence of alleles advantageous in the new environments. Heritable variations that alter the susceptibility to diseases vary with the historical period, the virulence of the infections, and their geographical spread. In this study we built polygenic scores for heritable traits that influence the genetic adaptation in the production of cytokines and immune-mediated disorders, including infectious, inflammatory, and autoimmune diseases, and applied them to the genomes of several ancient European populations. We observed that the advent of the Neolithic was a turning point for immune-mediated traits in Europeans, favoring those alleles linked with the development of tolerance against intracellular pathogens and promoting inflammatory responses against extracellular microbes. These evolutionary patterns are also associated with an increased presence of traits related to inflammatory and auto-immune diseases.
    • Therapeutic HNF4A mRNA attenuates liver fibrosis in a preclinical model.

      Yang, Taihua; Poenisch, Marion; Khanal, Rajendra; Hu, Qingluan; Dai, Zhen; Li, Ruomeng; Song, Guangqi; Yuan, Qinggong; Yao, Qunyan; Shen, Xizhong; et al. (Elsevier, 2021-08-25)
      Background & aims: Therapeutic targeting of injuries that require transient restoration of proteins by mRNA delivery is an attractive approach that, until recently, has remained poorly explored. In this study, we examined the therapeutic utility of mRNA delivery for liver fibrosis and cirrhosis. Specifically, we aimed to demonstrate the therapeutic efficacy of human hepatocyte nuclear factor alpha (HNF4A) mRNA in mouse models of fibrosis and cirrhosis. Methods: We investigated restoration of hepatocyte functions by HNF4A mRNA transfection in vitro, and analyzed the attenuation of liver fibrosis and cirrhosis in multiple mouse models, by delivering hepatocyte-targeted biodegradable lipid nanoparticles (LNPs) encapsulating HNF4A mRNA. To identify potential mechanisms of action, we performed microarray-based gene expression profiling, single-cell RNA sequencing, and chromatin immunoprecipitation. We used primary liver cells and human liver buds for additional functional validation. Results: Expression of HNF4A mRNA led to restoration of the metabolic activity of fibrotic primary murine and human hepatocytes in vitro. Repeated in vivo delivery of LNP-encapsulated HNF4A mRNA induced a robust inhibition of fibrogenesis in 4 independent mouse models of hepatotoxin- and cholestasis-induced liver fibrosis. Mechanistically, we discovered that paraoxonase 1 is a direct target of HNF4A and it contributes to HNF4A-mediated attenuation of liver fibrosis via modulation of liver macrophages and hepatic stellate cells. Conclusion: Collectively, our findings provide the first direct preclinical evidence of the applicability of HNF4A mRNA therapeutics for the treatment of fibrosis in the liver. Lay summary: Liver fibrosis and cirrhosis remain unmet medical needs and contribute to high mortality worldwide. Herein, we take advantage of a promising therapeutic approach to treat liver fibrosis and cirrhosis. We demonstrate that restoration of a key gene, HNF4A, via mRNA encapsulated in lipid nanoparticles decreased injury in multiple mouse models of fibrosis and cirrhosis. Our study provides proof-of-concept that mRNA therapy is a promising strategy for reversing liver fibrosis and cirrhosis.
    • Transient Depletion of Foxp3 Regulatory T Cells Selectively Promotes Aggressive β Cell Autoimmunity in Genetically Susceptible DEREG Mice.

      Watts, Deepika; Janßen, Marthe; Jaykar, Mangesh; Palmucci, Francesco; Weigelt, Marc; Petzold, Cathleen; Hommel, Angela; Sparwasser, Tim; Bonifacio, Ezio; Kretschmer, Karsten; et al. (Frontiers, 2021-08-10)
      Type 1 diabetes (T1D) represents a hallmark of the fatal multiorgan autoimmune syndrome affecting humans with abrogated Foxp3+ regulatory T (Treg) cell function due to Foxp3 gene mutations, but whether the loss of Foxp3+ Treg cell activity is indeed sufficient to promote β cell autoimmunity requires further scrutiny. As opposed to human Treg cell deficiency, β cell autoimmunity has not been observed in non-autoimmune-prone mice with constitutive Foxp3 deficiency or after diphtheria toxin receptor (DTR)-mediated ablation of Foxp3+ Treg cells. In the spontaneous nonobese diabetic (NOD) mouse model of T1D, constitutive Foxp3 deficiency did not result in invasive insulitis and hyperglycemia, and previous studies on Foxp3+ Treg cell ablation focused on Foxp3DTR NOD mice, in which expression of a transgenic BDC2.5 T cell receptor (TCR) restricted the CD4+ TCR repertoire to a single diabetogenic specificity. Here we revisited the effect of acute Foxp3+ Treg cell ablation on β cell autoimmunity in NOD mice in the context of a polyclonal TCR repertoire. For this, we took advantage of the well-established DTR/GFP transgene of DEREG mice, which allows for specific ablation of Foxp3+ Treg cells without promoting catastrophic autoimmune diseases. We show that the transient loss of Foxp3+ Treg cells in prediabetic NOD.DEREG mice is sufficient to precipitate severe insulitis and persistent hyperglycemia within 5 days after DT administration. Importantly, DT-treated NOD.DEREG mice preserved many clinical features of spontaneous diabetes progression in the NOD model, including a prominent role of diabetogenic CD8+ T cells in terminal β cell destruction. Despite the severity of destructive β cell autoimmunity, anti-CD3 mAb therapy of DT-treated mice interfered with the progression to overt diabetes, indicating that the novel NOD.DEREG model can be exploited for preclinical studies on T1D under experimental conditions of synchronized, advanced β cell autoimmunity. Overall, our studies highlight the continuous requirement of Foxp3+ Treg cell activity for the control of genetically pre-installed autoimmune diabetes.
    • Recent Developments on the Synthesis and Bioactivity of Ilamycins/Rufomycins and Cyclomarins, Marine Cyclopeptides That Demonstrate Anti-Malaria and Anti-Tuberculosis Activity.

      Kazmaier, Uli; Junk, Lukas; HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany. (MDPI, 2021-08-03)
      Ilamycins/rufomycins and cyclomarins are marine cycloheptapeptides containing unusual amino acids. Produced by Streptomyces sp., these compounds show potent activity against a range of mycobacteria, including multidrug-resistant strains of Mycobacterium tuberculosis. The cyclomarins are also very potent inhibitors of Plasmodium falciparum. Biosynthetically the cyclopeptides are obtained via a heptamodular nonribosomal peptide synthetase (NRPS) that directly incorporates some of the nonproteinogenic amino acids. A wide range of derivatives can be obtained by fermentation, while bioengineering also allows the mutasynthesis of derivatives, especially cyclomarins. Other derivatives are accessible by semisynthesis or total syntheses, reported for both natural product classes. The anti-tuberculosis (anti-TB) activity results from the binding of the peptides to the N-terminal domain (NTD) of the bacterial protease-associated unfoldase ClpC1, causing cell death by the uncontrolled proteolytic activity of this enzyme. Diadenosine triphosphate hydrolase (PfAp3Aase) was found to be the active target of the cyclomarins in Plasmodia. SAR studies with natural and synthetic derivatives on ilamycins/rufomycins and cyclomarins indicate which parts of the molecules can be simplified or otherwise modified without losing activity for either target. This review examines all aspects of the research conducted in the syntheses of these interesting cyclopeptides.
    • MicroRNA-125b-5p Regulates Hepatocyte Proliferation During the Termination Phase of Liver Regeneration.

      Yang, Dakai; Dai, Zhen; Yang, Taihua; Balakrishnan, Asha; Yuan, Qinggong; Vondran, Florian W R; Manns, Michael P; Ott, Michael; Cantz, Tobias; Sharma, Amar Deep; et al. (Wiley, 2020-09-15)
      The ability of the liver to regenerate and restore mass limits the increasing mortality rate due to life-threatening liver diseases. Successful liver regeneration is accomplished in multiple stages, of which the priming and proliferation phases are well studied. However, the regulatory pathways, specifically microRNA (miRNA)-mediated posttranscriptional regulation, which prevent uncontrolled proliferation and mediate the termination of liver regeneration, are not well understood. We identified differentially regulated miRNAs during the termination phase after 2/3 partial hepatectomy (PH) in mice, which is a well-established mouse model of liver regeneration. We further evaluated the function of differentially regulated miRNAs in primary mouse hepatocytes by using mimics and inhibitors and in vivo by using adeno-associated virus (AAV) serotype 8. A candidate miRNA target was identified by messenger RNA array in silico analyses and validated in primary mouse and human hepatocytes. Using miRNA profiling, we discovered miR-125b-5p as a novel regulator of hepatocyte proliferation in the late phase of liver regeneration. AAV-mediated miR-125b-5p delivery in mice enhanced the endogenous regenerative capacity and resulted in improved restoration of liver mass after 2/3 PH. Further, we found that ankyrin repeat and BTB/POZ domain containing protein 1 (Abtb1) is a direct target of miR-125b-5p in primary mouse and human hepatocytes and contributes to the pro-proliferative activity of miR-125b-5p by forkhead box G1 (FOXG1) and the cyclin-dependent kinase inhibitor 1A (p21) pathway. Conclusion: miR-125b-5p has an important role in regulating hepatocyte proliferation in the termination phase of liver regeneration and may serve as a potential therapeutic target in various liver diseases that often exhibit deregulated hepatocyte proliferation.
    • Staphylococcus epidermidis Phages Transduce Antimicrobial Resistance Plasmids and Mobilize Chromosomal Islands.

      Fišarová, Lenka; Botka, Tibor; Du, Xin; Mašlaňová, Ivana; Bárdy, Pavol; Pantůček, Roman; Benešík, Martin; Roudnický, Pavel; Winstel, Volker; Larsen, Jesper; et al. (American Society of Microbiology, 2021-05-12)
      Staphylococcus epidermidis is a leading opportunistic pathogen causing nosocomial infections that is notable for its ability to form a biofilm and for its high rates of antibiotic resistance. It serves as a reservoir of multiple antimicrobial resistance genes that spread among the staphylococcal population by horizontal gene transfer such as transduction. While phage-mediated transduction is well studied in Staphylococcus aureus, S. epidermidis transducing phages have not been described in detail yet. Here, we report the characteristics of four phages, 27, 48, 456, and 459, previously used for S. epidermidis phage typing, and the newly isolated phage E72, from a clinical S. epidermidis strain. The phages, classified in the family Siphoviridae and genus Phietavirus, exhibited an S. epidermidis-specific host range, and together they infected 49% of the 35 strains tested. A whole-genome comparison revealed evolutionary relatedness to transducing S. aureus phietaviruses. In accordance with this, all the tested phages were capable of transduction with high frequencies up to 10-4 among S. epidermidis strains from different clonal complexes. Plasmids with sizes from 4 to 19 kb encoding resistance to streptomycin, tetracycline, and chloramphenicol were transferred. We provide here the first evidence of a phage-inducible chromosomal island transfer in S. epidermidis Similarly to S. aureus pathogenicity islands, the transfer was accompanied by phage capsid remodeling; however, the interfering protein encoded by the island was distinct. Our findings underline the role of S. epidermidis temperate phages in the evolution of S. epidermidis strains by horizontal gene transfer, which can also be utilized for S. epidermidis genetic studies.IMPORTANCE Multidrug-resistant strains of S. epidermidis emerge in both nosocomial and livestock environments as the most important pathogens among coagulase-negative staphylococcal species. The study of transduction by phages is essential to understanding how virulence and antimicrobial resistance genes spread in originally commensal bacterial populations. In this work, we provide a detailed description of transducing S. epidermidis phages. The high transduction frequencies of antimicrobial resistance plasmids and the first evidence of chromosomal island transfer emphasize the decisive role of S. epidermidis phages in attaining a higher pathogenic potential of host strains. To date, such importance has been attributed only to S. aureus phages, not to those of coagulase-negative staphylococci. This study also proved that the described transducing bacteriophages represent valuable genetic modification tools in S. epidermidis strains where other methods for gene transfer fail.
    • Integration of metabolomics, genomics, and immune phenotypes reveals the causal roles of metabolites in disease.

      Chu, Xiaojing; Jaeger, Martin; Beumer, Joep; Bakker, Olivier B; Aguirre-Gamboa, Raul; Oosting, Marije; Smeekens, Sanne P; Moorlag, Simone; Mourits, Vera P; Koeken, Valerie A C M; et al. (BMC, 2021-07-06)
      Background: Recent studies highlight the role of metabolites in immune diseases, but it remains unknown how much of this effect is driven by genetic and non-genetic host factors. Result: We systematically investigate circulating metabolites in a cohort of 500 healthy subjects (500FG) in whom immune function and activity are deeply measured and whose genetics are profiled. Our data reveal that several major metabolic pathways, including the alanine/glutamate pathway and the arachidonic acid pathway, have a strong impact on cytokine production in response to ex vivo stimulation. We also examine the genetic regulation of metabolites associated with immune phenotypes through genome-wide association analysis and identify 29 significant loci, including eight novel independent loci. Of these, one locus (rs174584-FADS2) associated with arachidonic acid metabolism is causally associated with Crohn's disease, suggesting it is a potential therapeutic target. Conclusion: This study provides a comprehensive map of the integration between the blood metabolome and immune phenotypes, reveals novel genetic factors that regulate blood metabolite concentrations, and proposes an integrative approach for identifying new disease treatment targets.
    • Lactate dehydrogenase B regulates macrophage metabolism in the tumor microenvironment.

      Frank, Ann-Christin; Raue, Rebecca; Fuhrmann, Dominik C; Sirait-Fischer, Evelyn; Reuse, Carsten; Weigert, Andreas; Lütjohann, Dieter; Hiller, Karsten; Syed, Shahzad Nawaz; Brüne, Bernhard; et al. (Ivyspring International publisher, 2021-06-04)
      Background: Glucose metabolism in the tumor-microenvironment is a fundamental hallmark for tumor growth and intervention therein remains an attractive option for anti-tumor therapy. Whether tumor-derived factors such as microRNAs (miRs) regulate glucose metabolism in stromal cells, especially in tumor-associated macrophages (TAMs), to hijack them for trophic support, remains elusive. Methods: Ago-RIP-Seq identified macrophage lactate dehydrogenase B (LDHB) as a target of tumor-derived miR-375 in both 2D/3D cocultures and in murine TAMs from a xenograft mouse model. The prognostic value was analyzed by ISH and multiplex IHC of breast cancer patient tissues. Functional consequences of the miR-375-LDHB axis in TAMs were investigated upon mimic/antagomir treatment by live metabolic flux assays, GC/MS, qPCR, Western blot, lentiviral knockdown and FACS. The therapeutic potential of a combinatorial miR-375-decoy/simvastatin treatment was validated by live cell imaging. Results: Macrophage LDHB decreased in murine and human breast carcinoma. LDHB downregulation increase aerobic glycolysis and lactagenesis in TAMs in response to tumor-derived miR-375. Lactagenesis reduced fatty acid synthesis but activated SREBP2, which enhanced cholesterol biosynthesis in macrophages. LDHB downregulation skewed TAMs to function as a lactate and sterol/oxysterol source for the proliferation of tumor cells. Restoring of LDHB expression potentiated inhibitory effects of simvastatin on tumor cell proliferation. Conclusion: Our findings identified a crucial role of LDHB in macrophages and established tumor-derived miR-375 as a novel regulator of macrophage metabolism in breast cancer, which might pave the way for strategies of combinatorial cancer cell/stroma cell interventions.
    • Structure-Activity Relationship and Mode-of-Action Studies Highlight 1-(4-Biphenylylmethyl)-1H-imidazole-Derived Small Molecules as Potent CYP121 Inhibitors.

      Walter, Isabell; Adam, Sebastian; Gentilini, Maria Virginia; Kany, Andreas M; Brengel, Christian; Thomann, Andreas; Sparwasser, Tim; Köhnke, Jesko; Hartmann, Rolf W; HIPS, Helmholtz-Institut für Pharmazeutische Forschung Saarland, Universitätscampus E8.1 66123 Saarbrücken, Germany.; TWINCORE, Zentrum für experimentelle und klinische Infektionsforschung GmbH,Feodor-Lynen Str. 7, 30625 Hannover, Germany. (Wiley-VCH, 2021-05-19)
      CYP121 of Mycobacterium tuberculosis (Mtb) is an essential target for the development of novel potent drugs against tuberculosis (TB). Besides known antifungal azoles, further compounds of the azole class were recently identified as CYP121 inhibitors with antimycobacterial activity. Herein, we report the screening of a similarity-oriented library based on the former hit compound, the evaluation of affinity toward CYP121, and activity against M. bovis BCG. The results enabled a comprehensive SAR study, which was extended through the synthesis of promising compounds and led to the identification of favorable features for affinity and/or activity and hit compounds with 2.7-fold improved potency. Mode of action studies show that the hit compounds inhibit substrate conversion and highlighted CYP121 as the main antimycobacterial target of our compounds. Exemplified complex crystal structures of CYP121 with three inhibitors reveal a common binding site. Engaging in both hydrophobic interactions as well as hydrogen bonding to the sixth iron ligand, our compounds block a solvent channel leading to the active site heme. Additionally, we report the first CYP inhibitors that are able to reduce the intracellular replication of M. bovis BCG in macrophages, emphasizing their potential as future drug candidates against TB.
    • Care of patients with liver disease during the COVID-19 pandemic: EASL-ESCMID position paper.

      Boettler, Tobias; Newsome, Philip N; Mondelli, Mario U; Maticic, Mojca; Cordero, Elisa; Cornberg, Markus; Berg, Thomas; CiiM, Zentrum für individualisierte Infektionsmedizin, Feodor-Lynen-Str.7, 30625 Hannover. (Elsevier, 2020-04-02)
      The coronavirus disease 2019 (COVID-19) pandemic poses an enormous challenge to healthcare systems in affected communities. Older patients and those with pre-existing medical conditions have been identified as populations at risk of a severe disease course. It remains unclear at this point to what extent chronic liver diseases should be considered as risk factors, due to a shortage of appropriate studies. However, patients with advanced liver disease and those after liver transplantation represent vulnerable patient cohorts with an increased risk of infection and/or a severe course of COVID-19. In addition, the current pandemic requires unusual allocation of healthcare resources which may negatively impact the care of patients with chronic liver disease that continue to require medical attention. Thus, the challenge hepatologists are facing is to promote telemedicine in the outpatient setting, prioritise outpatient contacts, avoid nosocomial dissemination of the virus to patients and healthcare providers, and at the same time maintain standard care for patients who require immediate medical attention.
    • Cytotoxicity, Intracellular Replication, and Contact-Dependent Pore Formation of Genotyped Environmental Isolates from Hospital Water Systems in the West Bank, Palestine.

      Zayed, Ashraf R; Pecellin, Marina; Jaber, Lina; Butmeh, Suha; Bahader, Shereen A; Steinert, Michael; Höfle, Manfred G; Brettar, Ingrid; Bitar, Dina M; HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany. (MDPI, 2021-04-01)
      Legionella pneumophila is the causative agent of Legionnaires' disease. Due to the hot climate and intermittent water supply, the West Bank, Palestine, can be considered a high-risk area for this often fatal atypical pneumonia. L. pneumophila occurs in biofilms of natural and man-made freshwater environments, where it infects and replicates intracellularly within protozoa. To correlate the genetic diversity of the bacteria in the environment with their virulence properties for protozoan and mammalian host cells, 60 genotyped isolates from hospital water systems in the West Bank were analyzed. The L. pneumophila isolates were previously genotyped by high resolution Multi Locus Variable Number of Tandem Repeat Analysis (MLVA-8(12)) and sorted according to their relationship in clonal complexes (VACC). Strains of relevant genotypes and VACCs were compared according to their capacity to infect Acanthamoeba castellanii and THP-1 macrophages, and to mediate pore-forming cytotoxicity in sheep red blood cells (sRBCs). Based on a previous detailed analysis of the biogeographic distribution and abundance of the MLVA-8(12)-genotypes, the focus of the study was on the most abundant L. pneumophila- genotypes Gt4(17), Gt6 (18) and Gt10(93) and the four relevant clonal complexes [VACC1, VACC2, VACC5 and VACC11]. The highly abundant genotypes Gt4(17) and Gt6(18) are affiliated with VACC1 and sequence type (ST)1 (comprising L. pneumophila str. Paris), and displayed seroroup (Sg)1. Isolates of these two genotypes exhibited significantly higher virulence potentials compared to other genotypes and clonal complexes in the West Bank. Endemic for the West Bank was the clonal complex VACC11 (affiliated with ST461) represented by three relevant genotypes that all displayed Sg6. These genotypes unique for the West Bank showed a lower infectivity and cytotoxicity compared to all other clonal complexes and their affiliated genotypes. Interestingly, the L. pneumophila serotypes ST1 and ST461 were previously identified by in situ-sequence based typing (SBT) as main causative agents of Legionnaires' disease (LD) in the West Bank at a comparable level. Overall, this study demonstrates the site-specific regional diversity of L. pneumophila genotypes in the West Bank and suggests that a combination of MLVA, cellular infection assays and hierarchical agglomerative cluster analysis allows an improved genotype-based risk assessment.
    • Direct conversion of porcine primary fibroblasts into hepatocyte-like cells.

      Fráguas-Eggenschwiler, Mariane; Eggenschwiler, Reto; Söllner, Jenny-Helena; Cortnumme, Leon; Vondran, Florian W R; Cantz, Tobias; Ott, Michael; Niemann, Heiner; HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany. (Nature Research, 2021-04-29)
      The pig is an important model organism for biomedical research, mainly due to its extensive genetic, physiological and anatomical similarities with humans. Until date, direct conversion of somatic cells into hepatocyte-like cells (iHeps) has only been achieved in rodents and human cells. Here, we employed lentiviral vectors to screen a panel of 12 hepatic transcription factors (TF) for their potential to convert porcine fibroblasts into hepatocyte-like cells. We demonstrate for the first time, hepatic conversion of porcine somatic cells by over-expression of CEBPα, FOXA1 and HNF4α2 (3TF-piHeps). Reprogrammed 3TF-piHeps display a hepatocyte-like morphology and show functional characteristics of hepatic cells, including albumin secretion, Dil-AcLDL uptake, storage of lipids and glycogen and activity of cytochrome P450 enzymes CYP1A2 and CYP2C33 (CYP2C9 in humans). Moreover, we show that markers of mature hepatocytes are highly expressed in 3TF-piHeps, while fibroblastic markers are reduced. We envision piHeps as useful cell sources for future studies on drug metabolism and toxicity as well as in vitro models for investigation of pig-to-human infectious diseases.
    • Plasma Metabolome Signature Indicative of Germline Status Independent of Cancer Incidence.

      Penkert, Judith; Märtens, Andre; Seifert, Martin; Auber, Bernd; Derlin, Katja; Hille-Betz, Ursula; Hörmann, Philipp; Klopp, Norman; Prokein, Jana; Schlicker, Lisa; et al. (Frontiers, 2021-04-07)
      Individuals carrying a pathogenic germline variant in the breast cancer predisposition gene BRCA1 (gBRCA1+) are prone to developing breast cancer. Apart from its well-known role in DNA repair, BRCA1 has been shown to powerfully impact cellular metabolism. While, in general, metabolic reprogramming was named a hallmark of cancer, disrupted metabolism has also been suggested to drive cancer cell evolution and malignant transformation by critically altering microenvironmental tissue integrity. Systemic metabolic effects induced by germline variants in cancer predisposition genes have been demonstrated before. Whether or not systemic metabolic alterations exist in gBRCA1+ individuals independent of cancer incidence has not been investigated yet. We therefore profiled the plasma metabolome of 72 gBRCA1+ women and 72 age-matched female controls, none of whom (carriers and non-carriers) had a prior cancer diagnosis and all of whom were cancer-free during the follow-up period. We detected one single metabolite, pyruvate, and two metabolite ratios involving pyruvate, lactate, and a metabolite of yet unknown structure, significantly altered between the two cohorts. A machine learning signature of metabolite ratios was able to correctly distinguish between gBRCA1+ and controls in ~82%. The results of this study point to innate systemic metabolic differences in gBRCA1+ women independent of cancer incidence and raise the question as to whether or not constitutional alterations in energy metabolism may be involved in the etiology of BRCA1-associated breast cancer.
    • NeutrobodyPlex-monitoring SARS-CoV-2 neutralizing immune responses using nanobodies.

      Wagner, Teresa R; Ostertag, Elena; Kaiser, Philipp D; Gramlich, Marius; Ruetalo, Natalia; Junker, Daniel; Haering, Julia; Traenkle, Bjoern; Becker, Matthias; Dulovic, Alex; et al. (EMBO Press, 2021-04-27)
      In light of the COVID-19 pandemic, there is an ongoing need for diagnostic tools to monitor the immune status of large patient cohorts and the effectiveness of vaccination campaigns. Here, we present 11 unique nanobodies (Nbs) specific for the SARS-CoV-2 spike receptor-binding domain (RBD), of which 8 Nbs potently inhibit the interaction of RBD with angiotensin-converting enzyme 2 (ACE2) as the major viral docking site. Following detailed epitope mapping and structural analysis, we select two inhibitory Nbs, one of which binds an epitope inside and one of which binds an epitope outside the RBD:ACE2 interface. Based on these, we generate a biparatopic nanobody (bipNb) with viral neutralization efficacy in the picomolar range. Using bipNb as a surrogate, we establish a competitive multiplex binding assay ("NeutrobodyPlex") for detailed analysis of the presence and performance of neutralizing RBD-binding antibodies in serum of convalescent or vaccinated patients. We demonstrate that NeutrobodyPlex enables high-throughput screening and detailed analysis of neutralizing immune responses in infected or vaccinated individuals, to monitor immune status or to guide vaccine design.
    • Conservation of the HBV RNA element epsilon in nackednaviruses reveals ancient origin of protein-primed reverse transcription.

      Beck, Jürgen; Seitz, Stefan; Lauber, Chris; Nassal, Michael; TWINCORE, Zentrum für experimentelle und klinische Infektionsforschung GmbH,Feodor-Lynen Str. 7, 30625 Hannover, Germany. (Academy of Sciences, 2021-03-30)
      Hepadnaviruses, with the human hepatitis B virus as prototype, are small, enveloped hepatotropic DNA viruses which replicate by reverse transcription of an RNA intermediate. Replication is initiated by a unique protein-priming mechanism whereby a hydroxy amino acid side chain of the terminal protein (TP) domain of the viral polymerase (P) is extended into a short DNA oligonucleotide, which subsequently serves as primer for first-strand synthesis. A key component in the priming of reverse transcription is the viral RNA element epsilon, which contains the replication origin and serves as a template for DNA primer synthesis. Here, we show that recently discovered non-enveloped fish viruses, termed nackednaviruses [C. Lauber et al., Cell Host Microbe 22, 387-399 (2017)], employ a fundamentally similar replication mechanism despite their huge phylogenetic distance and major differences in genome organization and viral lifestyle. In vitro cross-priming studies revealed that few strategic nucleotide substitutions in epsilon enable site-specific protein priming by heterologous P proteins, demonstrating that epsilon is functionally conserved since the two virus families diverged more than 400 Mya. In addition, other cis elements crucial for the hepadnavirus-typical replication of pregenomic RNA into relaxed circular double-stranded DNA were identified at conserved positions in the nackednavirus genomes. Hence, the replication mode of both hepadnaviruses and nackednaviruses was already established in their Paleozoic common ancestor, making it a truly ancient and evolutionary robust principle of genome replication that is more widespread than previously thought.
    • Chemically Engineered Immune Cell-Derived Microrobots and Biomimetic Nanoparticles: Emerging Biodiagnostic and Therapeutic Tools

      Jahromi, Leila Pourtalebi; Shahbazi, Mohammad Ali; Maleki, Aziz; Azadi, Amir; Santos, Hélder A.; HIPS, Helmholtz-Institut für Pharmazeutische Forschung Saarland, Universitätscampus E8.1 66123 Saarbrücken, Germany. (Wiley-VCH, 2021-01-01)
      Over the past decades, considerable attention has been dedicated to the exploitation of diverse immune cells as therapeutic and/or diagnostic cell‐based microrobots for hard‐to‐treat disorders. To date, a plethora of therapeutics based on alive immune cells, surface‐engineered immune cells, immunocytes’ cell membranes, leukocyte‐derived extracellular vesicles or exosomes, and artificial immune cells have been investigated and a few have been introduced into the market. These systems take advantage of the unique characteristics and functions of immune cells, including their presence in circulating blood and various tissues, complex crosstalk properties, high affinity to different self and foreign markers, unique potential of their on‐demand navigation and activity, production of a variety of chemokines/cytokines, as well as being cytotoxic in particular conditions. Here, the latest progress in the development of engineered therapeutics and diagnostics inspired by immune cells to ameliorate cancer, inflammatory conditions, autoimmune diseases, neurodegenerative disorders, cardiovascular complications, and infectious diseases is reviewed, and finally, the perspective for their clinical application is delineated.
    • Clarithromycin impairs tissue-resident memory and Th17 responses to macrolide-resistant Streptococcus pneumoniae infections.

      Lindenberg, Marc; Almeida, Luis; Dhillon-LaBrooy, Ayesha; Siegel, Ekkehard; Henriques-Normark, Birgitta; Sparwasser, Tim; TWINCORE, Zentrum für experimentelle und klinische Infektionsforschung GmbH,Feodor-Lynen Str. 7, 30625 Hannover, Germany. (Springer, 2021-02-17)
      The increasing prevalence of antimicrobial resistance in pathogens is a growing public health concern, with the potential to compromise the success of infectious disease treatments in the future. Particularly, the number of infections by macrolide antibiotics-resistant Streptococcus pneumoniae is increasing. We show here that Clarithromycin impairs both the frequencies and number of interleukin (IL)-17 producing T helper (Th) 17 cells within the lungs of mice infected with a macrolide-resistant S. pneumoniae serotype 15A strain. Subsequently, the tissue-resident memory CD4+ T cell (Trm) response to a consecutive S. pneumoniae infection was impaired. The number of lung resident IL-17+ CD69+ Trm was diminished upon Clarithromycin treatment during reinfection. Mechanistically, Clarithromycin attenuated phosphorylation of the p90-S6-kinase as part of the ERK pathway in Th17 cells. Moreover, a strong increase in the mitochondrial-mediated maximal respiratory capacity was observed, while mitochondrial protein translation and mTOR sisgnaling were unimpaired. Therefore, treatment with macrolide antibiotics may favor the spread of antimicrobial-resistant pathogens not only by applying a selection pressure but also by decreasing the natural T cell immune response. Clinical administration of macrolide antibiotics as standard therapy procedure during initial hospitalization should be reconsidered accordingly and possibly be withheld until microbial resistance is determined. KEY MESSAGES: • Macrolide-resistant S. pneumoniae infection undergoes immunomodulation by Clarithromycin • Clarithromycin treatment hinders Th17 and tissue-resident memory responses • Macrolide antibiotics impair Th17 differentiation in vitro by ERK-pathway inhibition.
    • Selective Host Cell Death by Staphylococcus aureus : A Strategy for Bacterial Persistence.

      Missiakas, Dominique; Winstel, Volker; TWINCORE, Zentrum für experimentelle und klinische Infektionsforschung GmbH,Feodor-Lynen Str. 7, 30625 Hannover, Germany. (Frontiers, 2021-01-21)
      Host cell death programs are fundamental processes that shape cellular homeostasis, embryonic development, and tissue regeneration. Death signaling and downstream host cell responses are not only critical to guide mammalian development, they often act as terminal responses to invading pathogens. Here, we briefly review and contrast how invading pathogens and specifically Staphylococcus aureus manipulate apoptotic, necroptotic, and pyroptotic cell death modes to establish infection. Rather than invading host cells, S. aureus subverts these cells to produce diffusible molecules that cause death of neighboring hematopoietic cells and thus shapes an immune environment conducive to persistence. The exploitation of cell death pathways by S. aureus is yet another virulence strategy that must be juxtaposed to mechanisms of immune evasion, autophagy escape, and tolerance to intracellular killing, and brings us closer to the true portrait of this pathogen for the design of effective therapeutics and intervention strategies.
    • Regulatory T Cells in an Endogenous Mouse Lymphoma Recognize Specific Antigen Peptides and Contribute to Immune Escape.

      Ahmetlić, Fatima; Riedel, Tanja; Hömberg, Nadine; Bauer, Vera; Trautwein, Nico; Geishauser, Albert; Sparwasser, Tim; Stevanović, Stefan; Röcken, Martin; Mocikat, Ralph; et al. (American Association for Cancer Research (AACR), 2019-03-20)
      Foxp3+ regulatory T cells (Tregs) sustain immune homeostasis and may contribute to immune escape in malignant disease. As a prerequisite for developing immunologic approaches in cancer therapy, it is necessary to understand the ontogeny and the antigenic specificities of tumor-infiltrating Tregs. We addressed this question by using a λ-MYC transgenic mouse model of endogenously arising B-cell lymphoma, which mirrors key features of human Burkitt lymphoma. We show that Foxp3+ Tregs suppress antitumor responses in endogenous lymphoma. Ablation of Foxp3+ Tregs significantly delayed tumor development. The ratio of Treg to effector T cells was elevated in growing tumors, which could be ascribed to differential proliferation. The Tregs detected were mainly natural Tregs that apparently recognized self-antigens. We identified MHC class II-restricted nonmutated self-epitopes, which were more prevalent in lymphoma than in normal B cells and could be recognized by Tregs. These epitopes were derived from proteins that are associated with cellular processes related to malignancy and may be overexpressed in the tumor.
    • Misinterpretation of the odds ratios.

      Fernández, Nathalie; HZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany. (Elsevier, 2020-05-11)
      No abstract available